1. Field of the Invention
This invention relates to a prophylactic and therapeutic composition for inhibiting rotavirus infection, and more particularly to a method of preventing and treating diseases or conditions associated with, or requiring, rotavirus infection, using defatted human milk globules (HMFGs), milk macromolecular fraction, curd, whey, the human milk mucin complex or a polypeptide having the rotavirus-binding specificity of the 46 Kdalton (Kd) app. MW HMFG glycoprotein. The present method finds its application in the prevention and treatment of diseases such as infantile gastroenteritis, and diarrheal conditions that afflict immunodeficient patients, the elderly and travelers.
2. Description of the Background
Gastroenteritis and diarrhea have been linked to rotavirus infection in a variety of clinical settings. In many cases the population afflicted by these diseases are the very young, the elderly and the immunocompromised. Acute infectious gastrointestinal diseases, for example, are a major cause of illness and death in infants and young children throughout the world. In the developing countries, infectious gastrointestinal diseases are estimated to cause up to 12,000 deaths per day. Diarrheal disease is also an important health problem in the developed countries. In the U.S., over 200,000 children under 5 years of age are hospitalized each year with acute diarrheal disease. This results in nearly 880,000 in-patient hospital days, over 500 deaths, and almost one billion dollars of in-patient costs per year.
Although various viral, bacterial, and parasitic agents are suspected of causing acute infectious gastroenteritis, rotaviruses have been identified as the most important viral agent of gastroenteritis, e.g., in children living in both developed and developing countries. Prospective studies indicate that in the U.S. rotaviruses account for around 2.9 million yearly episodes of diarrhea leading to 22,000 annual hospitalizations of children less than 5 years old. Rotaviruses have also been implicated as the causative agent of diarrheal outbreaks occurring in nursing homes, day care centers and during travel, and resulting from adult contacts with sick children. Additionally, rotaviruses have been linked with the occurrence of diarrhea in patients undergoing bone marrow transplants and suffering from various immunodeficient conditions.
Throughout history, breast-feeding infants were shown to be somewhat protected against enteric infection by pathogens in general when compared with bottle fed infants. In addition, breast-feeding was also shown to lower the incidence of enteric diseases, such as necrotizing enterocolitis of infancy, which may be infectious in origin but which have not been associated with a single etiologic agent. More recently, studies of children living in developing as well as developed countries such as Great Britain and the U.S. have shown that breast-fed infants undergo substantially fewer episodes of gastroenteritis than bottle-fed infants. In some studies breast-feeding was shown to lessen the severity of diarrhea and vomiting associated with rotavirus infection in hospitalized children, but not to provide total protection against infection in general, and serious episodes of rotavirus infection in particular. However, no single factor was found to be responsible for this effect in spite of the fact that antibodies were suspected of being involved in the effect. The level of anti-rotavirus antibody in human milk was found not to correlate with the degree of protection afforded by the milk. This suggested that non-immunoglobulin factors may play a role in the protective process. Among the nonimmunoglobulin factors that have been implicated in this phenomenon are lipids, .alpha.-interferon and trypsin inhibitors, among others. It has also been suggested that some of these substances are possible inhibitors of viral replication and of microorganisms in general. A factor isolated from a milk fraction free of fat and cells was shown to inhibit the infectivity of respiratory syncytial virus (RSV). The factor has a molecular weight greater than 400 Kd app. MW, and is distinct from the present agents.
The growth of rotavirus and its infectivity were studied in various systems. Rotavirus replication was shown recently to be inhibited by avian egg and bovine submaxillary gland glycoproteins in cell culture. These glycoproteins bind to the virus and their activity requires sialic acid and proceeds by interference with the binding of the virus to cellular receptors. Intestinal brush border membranes were also shown to bind rotavirus by attachment to glycoproteins. Human milk fat globules (HMFG) are obtained from the cream fractions of milk, and have been utilized to prepare polyclonal and monoclonal antibodies for use in the diagnosis of breast cancer. Both, anti-HMFG and anti-breast tumor monoclonal antibodies with specificities for different epitopes of the mucin complex have been produced. The anti-HMFG monoclonal antibodies were used to identify a large molecular weight mucin-like complex called non-penetrating glycoprotein (NPGP) on the surface of breast epithelial cells. The human milk mucin is a highly glycosylated macromolecular complex consisting of 50% carbohydrate, most of which is O-linked. In addition to the mucin molecule, this complex contains a disulfide-linked 70 Kd apparent molecular weight (app. MW) glycoprotein and a 46 Kd app. MW glycoprotein. Monoclonal antibodies raised against the 70 Kd app.MW and 46 Kd app.MW glycoproteins have also been produced. The 46 Kd app. MW and 70 Kd app. MW glycoproteins are found in the serum of breast cancer patients and may thus be used as markers for breast cancer. The 70 Kd app.MW glycoprotein, in particular, was found to co-purify with the intact mucin complex and to be linked to the mucin complex through disulfide bonds, making it a suspect linker protein of this complex on the breast epithelial surface.
The structure of the polypeptide associated with the 70 Kd app.MW glycoprotein was determined by cDNA cloning. A partial amino acid sequence of the 70 Kd app. MW polypeptide has been reported. (Larocca, D. D., et al., Cancer Research 50:5925-5930 (1990)). The 46 Kd app.MW glycoprotein and its immune complexes were detected in the serum of breast cancer patients using monoclonal antibodies against the glycoprotein. In addition, an increase in the levels of the 46 Kd app.MW glycoprotein in the patients'serum was also found to be associated with the advent of tumors. The structure of the 46 Kd app.MW glycoprotein and the amino acid sequence of its polypeptide have been described. Also known are the anti-neoplastic and diagnostic use of this glycoprotein and its polypeptide as well as the corresponding DNA and RNA sequences of the polypeptide. (Larocca, D. D., et al., Cancer Research 51:4944-4998 ( 1991 )), the text of which relating to the preparation and characterization of the 46 Kd app.MW glycoprotein is incorporated herein by reference.
The recognition of the importance of rotaviral infection, and of its epidermiological and economic consequences, has led to a substantial effort directed at its prevention by means of active immunization. However, current vaccine regimens have displayed poor efficacy.
Accordingly, there still is a need for a potent and effective treatment for rotaviral infection as well as its prophylaxis, substantially lacking detrimental side effects.